The invention discloses a 3D printing pre-stressed floor slab. The 3D printing pre-stressed floor slab includes a 3D printing frame and pre-stressed reinforcing steel bars which are arranged in the 3D printing frame. The 3D printing pre-stressed floor slab is characterized by further comprising sleeves, the sleeves are arranged at the positions, where pre-stressing force needs to be applied, of the floor slab, longitudinally penetrate into the 3D printing frame and are fixed by means of a plurality of U-shaped steel bars arranged in the frame in advance, the pre-stressed reinforcing steel bars are arranged in the sleeves, and the two ends of each pre-stressed reinforcing steel bar are provided with anchoring parts. The invention further provides a construction method of the 3D printing pre-stressed floor slab, the pre-stressed reinforcing steel bars are longitudinally arranged in the frame, concrete is poured, the integrity of the floor slab is ensured, and the tensile strength of the 3D printing pre-stressed floor slab is increased. According to the 3D printing pre-stressed floor slab and the construction method thereof, the structure is reasonable, construction is convenient, the steel bar distribution problem of the 3D printing floor slab is solved, the integrity of the floor slab is ensured and the tensile strength of floor slab is improved.